Composition using cross-linked hyaluronic acid for topical cosmetic and therapeutic applications

ABSTRACT

Disclosed are compositions comprising crosslinked hyaluronic acid gels, preferably vinyl sulfone cross-linked hyaluronic acid known as hylan B gel, for use in topical cosmetic and dermatological formulations. The hylan B gel in these formulations provides prolonged delivery of incorporated substances to the surface of the skin, to provide a hydrated film on the surface of the skin, and to provide a substantive and compatible film on the skin.

BACKGROUND OF THE INVENTION Field of the Invention

The present invention relates to compositions containing cross-linkedhyaluronic acid (HA) gels including, but not limited to, hylan B gel,for use in topical cosmetic and dermatological applications.

Description of Related Art

Topical skin care product compositions often contain a variety ofnatural and/or synthetic polymeric substances, for the purpose ofdelivering active ingredients to the surface of the skin and/orproviding moisturization of the skin. For example, U.S. Pat. No.5,911,980 describes the use of lipophilic and amphiphilic or hydrophilicfilm-forming polymer compositions as a topical agent for the delivery ofagents to the skin.

U.S. Pat. No. 6,824,785 discloses multi-compartment or multi-layeredstructures that may be loaded with an active ingredient for dermaldelivery. Lipids, typically in the form of liposomes are used to providean epidermal barrier when applied topically to the skin, which reducestrans-epidermal water loss.

U.S. Pat. No. 6,719,740 describes a delivery system, comprised of a toplayer and a bottom layer attached to each other, with a pouch thatcontains the skin care ingredient between the two layers.

U.S. Pat. No. 6,746,689 describes method for administering an activeingredient where a local anesthetic is combined with an intradermalpenetration agent.

U.S. Pat. No. 6,316,011 describes a reverse thermally viscosifyingcomposition including a linear block copolymer, wherein at least oneblock comprises a polyoxamer having a hydrophobic region and ahydrophilic region effective to form micelles in response to a change intemperature, and at least one block comprises a bioadhesive polymer oroligomer in an aqueous medium for the administration of a pharmaceuticalagent to the skin.

U.S. Pat. No. 6,495,149 describes the use of a liquid carrier fordelivery of an active comprised of cyclomethicone liquid for topical,leave-on compositions.

U.S. Pat. No. 6,235,312 describes the use of liquid crystalline phasedrug delivery systems for topical delivery to the skin, wherein theliquid crystalline phase is comprised of a mono glyceride.

Several patents disclose compositions containing cross-linked hyaluronicacid gels for use in topical formulations. For example, U.S. Pat. No.4,582,865 describes cross-linked gels of hyaluronic acid or mixedcross-linked gels of hyaluronic acid, wherein the gels further compriseat least one other hydrophilic polymer having a functional group capableof reacting with divinyl sulfone and either an inert water insolublesubstance or a low molecular weight substance covalently bonded to themacromolecular network, and methods of making these gels.

U.S. Pat. No. 4,605,691 describes methods of preparing a cross-linkedgel of hyaluronic acid comprising subjecting sodium hyaluronate in adilute aqueous alkaline solution at a pH of not less than about 9 to across-linking reaction with divinyl sulfone at about 20° C. and across-linked gel of hyaluronic acid produced by this method.

U.S. Pat. No. 4,636,524 describes a delivery system for a substancehaving biological or pharmacological activity comprising a molecularcage formed of a cross-linked gel of hyaluronic acid or a mixedcross-linked gel of hyaluronic acid and at least one other hydrophilicpolymer and containing a diffusible substance having biological orpharmacological activity.

U.S. Pat. No. 5,143,724 describes biocompatible gel slurries consistingof two phases: a particulate gel phase comprised of a chemicallycross-linked polymeric gel and a second, fluid phase comprising apolymer solution of a water-soluble biocompatible polymer in the aqueousmedium, wherein the fluid phase has a pH in the range of 5.0-8.0.

U.S. Pat. No. 5,246,698 describes methods of making the slurries of U.S.Pat. No. 5,143,724.

U.S. Pat. No. 5,399,351 describes methods of using these slurries tocontrol adhesion formation between tissues of a living body resultingfrom surgical intervention.

European Patent 0224987 describes drug delivery systems based onhyaluronic acid and its salts and derivatives thereof in soluble ornon-soluble cross-linked forms, which serve as a vehicle for slowrelease of a drug from a system.

BRIEF SUMMARY OF THE INVENTION

The present invention, in one aspect thereof, provides compositionscomprising cross-linked hyaluronic acid (HA) gels including, but notlimited to, hylan B gel, for use in topical cosmetic and dermatologicalapplications. The hylan B gel acts as a hydrated molecular cage as thatterm is described in U.S. Pat. No. 4,636,524, and into which one mayincorporate and entrap substances, including water, for delivery thereofto the surface of the skin. Typically, the composition contains anactive ingredient at a concentration of 0-100%, preferably 0.1-50% andmost preferably, 10-20% by weight of the total composition.

In another aspect thereof, the invention provides a topical compositioncomprising (a) a polysaccharide gel matrix, in combination with (b)another substance, which may be (i) a hydrophilic organic ingredient,such as an α-, β-, γ- or δ-hydroxy acid, an α-amino acid, a β-aminoacid, a γ-amino acid, a δ-amino acid, a glycerin, a vitamin or theirderivatives, a sugar, an essential oil, a water-soluble oil, afragrance, a natural polymer, a synthetic polymer, a cross-linkednatural polymer, a cross-linked synthetic polymer, a peptide, a protein,a polysaccharide or a nucleic acid; wherein the concentration of thehydrophilic organic ingredient may be from 0-100%, preferably, 0.1-10%and most preferably, from 1-5% by weight based on the entire composition(ii) a hydrophobic organic ingredient, such as a petrolatum, a fattyacid, a fatty alcohol, an oil, a lipid or a silicone; wherein theconcentration of the hydrophobic organic ingredient may be from 0-100%,preferably 0.1-50% by weight based on the weight of the entirecomposition; (iii) an inorganic ingredient such as titanium dioxide,silica, calcium chloride, kaolin, talc or tantalum, alone, wherein theconcentration of the inorganic ingredient may be from 0-40%, preferably0.1-4% by weight based on the weight of the entire composition, saidinorganic ingredient being present alone, or in combination with, aneffective amount of a therapeutic agent deliverable to a skin surface,wherein the therapeutic agent is a salt, a solvate, a prodrug, and aderivative of the therapeutic agent, and wherein the substance, alone orin combination, is entrapped within the matrix. In this embodiment, thetherapeutic agent is an anti-inflammatory agent such as a steroid, forexample, hydrocortisone, hydrocortisone acetate or prednison, anantiseptic agent, such as iodine, an anti-infective agent, i.e., anantibiotic such as bacitracin, a mucous membrane agent, a cleansingagent, a preservative, an astringent, an antihistamine, an analgesic, ananesthetic, an acne medication, an antifungal agent, an anti-pruriticagent, a hormone, a growth factor, a moisturizing agent, a sunscreensuch as Parsol MCX, a hyperpigmentation agent, an antioxidant or anutritional agent which is a vitamin or a mineral. The amount of thetherapeutic agent present in the composition may be anywhere from 0% upto the specific recommended dosage for each such specific therapeuticagent. For example, in the case of an antibiotic such as bacitracin, thespecific dosage will typically be about 500 units per gram of thecomposition in the form of an ointment. In the case of a steroid, suchas hydrocortisone, the specified dosage will typically be about 0.25-1%by weight, and in the case of a therapeutic agent such as capsaicin, thespecified dosage will typically be in the amount of about 0.25% byweight of the entire composition.

In another embodiment, the therapeutic agent of the composition is amixture of the anti-inflammatory agent and the anti-infective agent.

In another embodiment, the invention provides a composition wherein thepolysaccharide of the composition is a cross-linked polymer ofhyaluronic acid, a salt of the polymer or a derivative of the polymer.

In yet another embodiment of the invention, the polymer is in anequilibrium or non-equilibrium form and is hylan B, where the hylan Bconcentration is from about 0.4-20% by weight.

BRIEF DESCRIPTION OF THE FIGURE

FIG. 1 illustrates higher levels of sunscreen on the surface of skin inthe presence of HY. 0.1 gm samples of each sunscreen composition wereweighed out onto weigh paper squares, and then each applied todesignated, marked areas on the washed forearm of human volunteers.After 2 hours each area was swabbed with a cotton tip applicator, thenthe site was stripped 8 times using Cuderm® stripping disks. Each diskwas placed into a test tube containing 3 ml of isopropanol, 2 disks wereplaced per tube, to elute the Parsol sunscreen from the skin on thedisk. The absorbance of the isopropanol in each tube was measured at λmax 311 nm). The data in the graph indicate that higher levels ofsunscreen are present on the uppermost layers of the skin, and thatthere is more sunscreen on the surface of the skin in the samplecontaining Hylasome (far left side of graph). Deeper layers of the skincontain less sunscreen, but show a trend toward more sunscreen beingabsorbed into the skin from the control (non-Hylasome) samples. SurfaceRetention of sunscreen at 2 hours is shown for four surfaces.

DETAILED DESCRIPTION OF THE INVENTION

The compositions of the present invention, which contain crosslinkedhyaluronic acid (HA) gels including, but not limited to, hylan B gel,are compositions for use in topical cosmetic and dermatologicalapplications and in which hylan B gel is used as a hydrated molecularcage into which one may incorporate, entrap and deliver substances,including water, to the surface of the skin. The hylan B gel may beeither in equilibrium or non-equilibrium form. In the non-equilibriumform, the gel is not fully swollen, while in the equilibrium form, thegel is swollen fully and cannot swell further. Hylan B gel matrix isderived from hyaluronic acid, otherwise known as hyaluronan or HA, andis a vinyl sulfone cross-linked derivative of hyaluronic acid. The hylanB gel matrix has an unusually high capacity to bind water, and inparticular, to a greater degree than other natural and syntheticpolymers [Takigami et al., Carbohydrate Polymers 26N: 11-18 (1995)].Water, which is a slightly bound portion of the gel matrix and whichforms reservoirs in which ingredients can be incorporated or trapped, iseffectively delivered to the skin because of its bound state.

The ingredients which are incorporated into, or entrapped, within thehylan B gel matrix (equilibrated or non-equilibrated) include, but arenot limited to, some classes of organic compounds, inorganic compounds,and mixtures thereof.

Among the organic compounds included are hydrophilic organic substancesincluding, but not limited to an α-, β-, γ- or δ-hydroxy acid, α-, β-,γ-, and δ-amino acids, glycerin, vitamins and their derivatives, sugars,essential oils, water soluble oils and fragrances, low molecular weightand high molecular weight substances including natural and syntheticpolymers, cross-linked natural and synthetic polymers, peptides,proteins, water soluble hydrophilic proteins, such as albumins,polysaccharides, nucleic acids, and mixtures thereof. The organiccompounds also include, but are not limited to, hydrophobic organicsubstances, including, but not limited to, petrolatum, fatty acids,fatty alcohols, oils, lipids, silicones, and mixtures thereof. Among theinorganic compounds and substances included are substances such as, butnot limited to, titanium dioxide, silica, calcium chloride, kaolin,talc, tantalum and mixtures thereof.

Ingredients included in these-chemical-categories include activeingredients, drugs, prodrugs and nondrugs. They include, but are notlimited to, the pharmaceutical categories of anti-inflammatory agents,antiseptics, anti-infectives, mucous membrane agents, cleansing agents,preservatives, astringents, antihistamines, analgesics, capsaicin, acnemedications, antifungals, antipruritics, hormones, growth factors,moisturizers, sunscreens, hyperpigmentation agents, antioxidants,nutritional substances, including, but not limited to, vitamins andminerals, substances of botanical, marine and animal origin, homeopathicsubstances and mixtures thereof. As should be clear from the foregoingdescription, very many different types of materials can be and arecapable of being incorporated into the hylan B gel for the purpose ofdelivering them to the skin of a host, typically, a human, but notnecessarily a human.

An advantage of this invention is the ability to provide prolonged andeffective delivery of incorporated ingredients to the surface of theskin and, where applicable to provide increased protection from theirritant effects of some active ingredients, for example, from theburning sensations produced by some active ingredients, e.g., capsaicin.Additionally, one can provide a hydrated film on the surface of the skinin which certain ingredients have been incorporated, and which containswater in the form of slightly bound water, and as well, to provide asubstantive and compatible film on the surface of the skin.

Hyaluronan or hyaluronic acid (HA) is a naturally occurringpolysaccharide that consists of alternating N-acetyl-D-glucosamine andD-glucuronic acid monosaccharide units linked with alternating [beta]1-3 glucoronidic (?) and [beta] 1-4 glucosaminidic (?) bonds. Themolecular weight of hyaluronic acid is generally within the range of5×10⁴ up to more than 8×10⁶ Daltons. Cross-linked HA gels include, butare not limited to, hylan B gel matrix, which is a vinyl sulfonecross-linked derivative of hyaluronic acid. Cross linked simple andmixed gels based on hyaluronan, products containing such gels, andmethods to prepare same are described in U.S. Pat. Nos. 4,582,865,4,605,691 and 4,636,524, which are noted above in the Description ofRelated Art, and the contents of which are incorporated by referenceherein. It should also be clearly understood that hyaluronan of anyorigin may be successfully used for the purposes of this invention.

The compositions or the present invention may be administered in dosageunit formulations containing conventional nontoxic pharmaceuticallyacceptable carriers, adjuvants, and vehicles as desired. Topicaladministration also may involve the use of transdermal administrationsuch as transdermal patches or iontophoresis devices which are preparedaccording to techniques and procedures well known in the art.

The amount of the compositions of the present invention which will beeffective in the treatment of a particular disorder or condition willdepend on the nature of the disorder or condition, and can be determinedby standard clinical techniques. See, for example, Goodman and Gilman;The Physician's Desk Reference, Medical Economics Company, Inc.,Oradell, N.J., 1995; and Drug Facts and Comparisons, Facts and,Comparisons, Inc., St. Louis, Mo., 1993. The precise dose to be employedin any particular formulation will also depend on the seriousness of thedisease or disorder, and should be decided upon and determined accordingto the judgment of the practitioner and each patient's specificcircumstances.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of ordinary skillin the art to which this invention pertains. Although any methods andmaterials similar or equivalent to those described herein can also beused in the practice or testing of the present invention, the preferredmethods and materials will now be described. All publications mentionedherein. are incorporated herein by reference to disclose and describethe methods and/or materials in connection with which the publicationsare cited. Unless indicated otherwise, parts are parts by weight,molecular weight is weight average molecular weight, temperature is indegrees Centigrade, and pressure is at or near atmospheric.

The term “effective amount” is used herein to denote that amount of anagent that will elicit the response of a mammal, including a human, thatis being sought for the purpose of effecting a desired therapeuticeffect. The term “therapeutically effective amount” is used herein todenote that amount of a drug or pharmaceutical agent or apharmaceutically acceptable salt, solvate or prodrug thereof, that willelicit the therapeutic response of an animal or human that is beingsought.

Where a range of values is provided, it is understood that eachintervening value, to the tenth of the unit of the lower limit, unlessthe context clearly dictates otherwise, between the upper and lowerlimit of that range and any other stated or intervening value in thatstated range is encompassed within the invention. The upper and lowerlimits of these smaller ranges which may independently be included inthe smaller ranges is also encompassed within the invention, subject toany specifically excluded limit in the stated range. Where the statedrange includes one or both of the limits, ranges excluding either bothof those included limits are also included in the invention.

The publications discussed herein are provided solely for theirdisclosure prior to the filing date of the present application. Nothingherein is to be construed as an admission that the present invention isnot entitled to antedate such publication by virtue of prior invention.Further, the dates of publication provided may be different from theactual publication dates which may need to be independently confirmed.

The invention has been described with reference to the preferredembodiments to illustrate the principles of the invention and not tolimit the invention to the particular embodiments illustrated.Modifications and alterations may occur to others upon reading andunderstanding the preceding detailed description. It is intended thatthe scope of the invention be construed as including all modificationsand alterations that may occur to others upon reading and understandingthe preceding detailed description insofar as they come within the scopeof the following claims or equivalents thereof.

The invention is described in more detail and with respect to only some,but not all embodiments thereof in the following examples, which are notintended to limit the scope of the claims.

As will be evident from the following examples, the properties of thetopical compositions of the present invention are superior to thosedescribed in the prior art mentioned above. The present compositions allform a permeable, hydrated film on the surface of the skin, require noenvironmental stimulus, such as temperature change, to deliver, sincedelivery of the active occurs upon application of the compositions tothe skin, and they suffer from little or no temperature sensitivity, andtherefore experience no marked changes in the physical state of thecomposition upon exposure to high and low ambient temperatures.

Example 1

This example illustrates the preparation of nonequilibrium gel ofhyaluronic acid (Hylasome a sodium hyaluronate cross-polymer) with apolymer concentration of approximately 1%. 30 gm of sodium hyaluronate(HA) of bacterial origin were dissolved in 853 gm of distilled water. Tothis solution 100 gm of 1N NaOH were added to achieve a finalconcentration of 0.1N alkali. The mixture was stirred carefully untiluniform. To this solution a mixture of 7 gm (6 ml) of divinyl sulfone(DVS) in 10 gm of water was added and stirred until a strong gel wasformed with an HA concentration of approximately 3%; typically 15-20minutes for completion of the cross-linking reaction. The resultant gel(approximately 1 kg) was placed into dialysis tubing (32 mm diameter,12000-14000 molecular weight cut-off, Spectrum) so that the gel occupiedapproximately one-third of the length of the dialysis tubing. The end ofthe dialysis tubing was sealed and the gel manually distributedthroughout the length of the tubing in order to facilitate uniformswelling. The sealed tubing containing gel was placed into a cleancontainer with five liters of purified water; the water was changedtwice daily until the pH of the gel inside the tubing was neutral (pH6-7). The neutral, washed gel was expelled from the dialysis tubing. Thegel was carefully mixed with a preservative (15 gm of phenonip dispersedin about 100 gm of water, to a final phenonip concentration of 0.5%)until uniform. The concentration of HA in the nonequilibrium gel wasapproximately 1% (10 mg/ml). The desigation “%” refers to theconcentration of HA in the gel, calculated based on the amount of dry HApolymer present in the gel.

Example 2: Hylasome (HA 0.1%)

This example illustrates the preparation of a swollen HA gel with apolymer concentration of 0.1%. To prepare 1 kg of this gel Hylasome (HA1%) in an amount of 100 gm was used. The Hylasome (HA 1%) was preparedaccording to Example 1. To 896 gm of RO (reverse osmosis) water wereadded 4 gm of phenonip-preservative and the mixture was stirred until auniform fine dispersion formed and then added to 100 gm of Hylasome (HA1%). The resultant swollen gel was then disintegrated using a high shearmixer (Silverson) using a low shear mixer blade, at a speed of 4000 rpm,for 4 minutes per 1 kg volume. The homogenized gel had a swelling ratioof 1,000. This material was used in some cosmetic and dermatologicalformulations as a highly effective hydrating component which slowlydelivers water. Hylasome (HA 0.1%) is a partially equilibrated gel andcan be further swollen with additional water and/or other activeingredients.

Example 3: Hylasome (HA 0.05%)

This example illustrates the preparation of 1 kg of fully swollenequilibrated gel with an HA concentration of 0.05%. This equilibratedgel can be made either from Hylasome (HA 1%) by adding 50 gm of this gelto 896 gm of RO water and 4 gm of phenonip preservative or by adding 500gm of Hylasome (HA 0.1%) to 500 gm of RO water and 2.23 gm of phenonip.The subsequent procedures are the same as described in Example 2.Hylasome (HA 1%) was prepared according to Example 1, and Hylasome (HA0.1%) was prepared according to Example 2. The resultant Hylasome (HA0.05%) has a swelling ratio of 2,000.

Example 4

This example illustrates the preparation of a formulation containingHylasome swollen with a solution of the sugars, glucose and mannitol.

Step 1: Preparation of the Active Ingredient—Hylasome Swollen with aSolution of Glucose and Mannitol.

To prepare 100 gm of this composition, to 62.6 gm of water was added 0.4gm of phenonip and mixed until a uniform dispersion formed. To thismixture were added 10.0 gm of D-mannitol and 13.5 gm of glucose. Thismixture was stirred until the sugars dissolved. Upon dissolution 13.5 gmof Hylasome (HA 1%) were added and the mixture stirred until the slurrywas uniform. The Hylasome gel slurry was then homogenized according tothe procedure described in Example 1. The concentration of sugars in thegel was 23.5%. The polymer concentration was 0.135%.

Parts Ingredients % (g/100 gm) 1A 1 Carbopol 940 0.36 2 water 53.67 3EDTA 0.14 4 Germaben II 0.75 1B 1 Cucumber extract 0.24 2 Aloe vera gel0.16 3 Chamomile extract 0.24 4 Glucam E-20 3.15 5 Apricot extract 0.246 Carrot extract 0.24 7 Tea extract 1.3 8 Eye bright extract 0.24 9 TEA(triethanolamine) 0.7 1C 1 Cellosize Polymer PCG019, 2% 4.72 with 5%alcohol 1.58 2 HA 1% 0.24 3 Polyox Coag 1% 0.48 4 Hydrosoy 2000 1D RoseNatural, water soluble 0.16 1E Dye - Orange/Red 2:1, 0.1% 0.12 Part 2 1Mannitol 2.97 Hylasome 2 Gluose 4.00 sugars 3 water 18.58 4 Hylasome4.00 5 Phenonip 0.12 TOTAL 29.67 Part 3 1 HA 1% 0.32 2 Hydrosoy 20000.16 3 DxS 0.004 4 Water 1.09 5 TEA (triethanolamine) 0.0006 6 phenonip0.16 TOTAL 1.73Step 2: Preparation of a Dermatological Formulation Containing HylasomeSwollen with a Solution of 23.5% Sugars.

This formulation was prepared as follows: Carbopol was dispersed inwater and the mixture was stirred until a uniform dispersion formed. Thepreservatives EDTA and Germaben II were then added. The ingredients inParts B and C were blended. Parts A and B were heated to 45-50° C. whilemixing separately, then combined together, stirred well and cooled to40° C. The components of Part C were blended and then heated to 40° C.and added to the combined Parts A and B, mixed carefully until uniformand then cooled to room temperature (20-25° C.). Part D was added andmixed, and then Part E was added and mixed by careful stirring. Thismixture constitutes Part 1. To Part 1 was added the Hylasome/sugarsslurry and after mixing, Part 3 was added. Part 3 was prepared byblending the listed ingredients. The pH of the composition was neutral(6-7). The sugar content was approximately 7%. This dermatologicalcomposition is used as a remedy for puffiness and minor swelling.

Example 5

Preparation of a face mask using Hylasome swollen with 47% lactic acidas active ingredient (INCI [International Nomenclature CosmeticIngredient] name sodium hyaluronate cross-polymer and lactic acid).

Step 1: Preparation of Hylasome Swollen with 47% Lactic Acid.

All ingredients in the following table except Hylasome were mixed. Whenmixing was complete, Hylasome was added and mixed until swollen. Themixture was homogenized until it became liquidy and uniform.

INGREDIENT %, gm/100 gm D-mannitol 2 Tris-amino 10 Glycerol 2.6 Lacticacid 47 Phenonip 0.4 RO water 18 Hylasome (HA 1%) 20Step 2: Preparation of the Basic Formulation, Containing the ActiveIngredient Hylasome Swollen with Lactic Acid.

This formulation was prepared in several stages, as follows: Part A1 wasprepared by dissolving pluronics (?) in water; Part A2 was made bydispersing Carbopol 940 in water and then adding the preservatives EDTAand Germaben II. Then Parts A1 and A2 were combined. The ingredients inPart B and C were blended; Parts A1, A2, and B were then heated to70-75° C. while mixing separately, then they were combined and mixingwas continued at 70-75° C. until uniform, followed by cooling to 45° C.Parts C and F were mixed, then combined and heated to 45° C., and addedto Parts A1+A2+B. This blend was cooled to room temperature, ca. 25° C.The pH of this mixture was neutral (6-7) adjusted by TEA (Part D). Tothis basic formulation was added the active ingredient Hylasome swollenwith 47% lactic acid (Part E). The pH of the formulation wasapproximately 3.5.

Part Ingredients % (g/100 g) A₁ 1 Water 35.57 2 Pluronic 87 1.90 3Pluronic 127 0.95 A₂ 1 Water 36.02 2 EDTA 0.14 3 Germaben II 0.50 4Carbopol 0.48 B 1 TEA (triethanolamine) 0.19 2 Tween 20 0.95 3Polydimethylsiloxane (fluid 200) 0.95 4 Phosal 50 PG 0.90 5 Beeswax 0.956 Robane (squalene) 1.43 7 Lanoxide -59 0.95 8 Coconut oil 0.48 9 Cetylalcohol 1.90 10 Propylene glycol 1.90 11 Glycerol monostearate 0.95 12Glucam E-20 1.90 13 Germaben II 0.50 14 Chamomile extract 0.19 15 LemonBioflavonoids extract 0.48 16 Calendula extract 0.19 17 Apricot fruitextract 0.10 18 Peppermint extract 0.05 C 1 HA 1% 0.95 2 Cellosize 2%(polymer PCG-10) 1.45 3 Jasmine/Rosewood Natural, water soluble 0.24 4Lavender Natural, water soluble 0.24 5 Lemon natural, water soluble 0.10D TEA (triethanolamine) 0.6 E Hylasome with 47% lactic acid (Step 2) 5.0F 1 HA 1% 0.19 2 Hydrosay 2000 0.095 3 DxS 0.0024 4 Water 0.60 5 TEA0.0033 6 Phenonip 0.0095

Step 3: Preparation of Face Mask

To the basic formulation containing the active component Hylasome-lacticacid were added 5 gm of ascorbic acid dispersed in 5 gm of Tween 20, and5 gm of water. To prepare 100 gm of the face mask, 83.3 gm of thiscomposition and 63 gm of titanium dioxide (TiO₂) which had been mixedwith 10.4 gm of Tween 20 were mixed. The pH of the face mask wasapproximately 3.0

Example 6

This example illustrates the preparation of an exfoliating cream withthe active ingredient Hylasome swollen with 34% glycolic acid.

Step 1: Preparation of the Active Ingredient (Hylasome-34% GlycolicAcid).

To prepare one (1) kg of this material, 20 gm of mannitol; 26 gm ofglycerol, 100 gm of tris-amino (buffer) and 340 gm of glycolic acid wereadded to 310 ml of RO water (reverse osmosis). These components werestirred together, then 4.0 gm phenonip (preservative) were added to thismixture. To the resulting mixture, 200 gm of Hylasome (HA 1%) were addedand the mixture stirred until fully swollen. The equilibrated gelcontained HA 0.2% and 34% glycolic acid, with a pH of approximately 3.0

Step 2: Preparation of Exfoliating Cream.

100 gm of exfoliating cream were made by adding 6 gm of the activeingredient complex (Hylasome-34% glycolic acid) to 94 gm of the basicformulation prepared according to Example 5 (Step 2).

Example 7

This example illustrates the preparation of compositions containingHylasome swollen with ascorbic acid (AA).

Step 1: Preparation of Compositions

Composition #1 Composition #2 Components (%, g/100 g) (%, g/100 g)Ascorbic Acid 9.4 9.4 Propylene glycol 58.0 55.3 glycerol 4.6 4.6 Ethylalcohol 4.6 4.6 Lemon bioflavinoids 0.46 0.46 mannitol — 2.7 Carbopol940 0.47 0.47 EDTA 0.10 0.10 Phenonip 0.50 0.50 TEA 1.3 1.3 Hylasome (HA1%) 10.2 10.2 Water (RO) 10.2 10.2 Natural lemon oil, water soluble 0.170.17

To prepare composition #1, water was added to Hylasome (HA 1%), in equalvolumes, and the gel allowed to swell with gentle mixing; the HAconcentration in the diluted Hylasome gel was 0.5%. To this gel wasadded dry powdered ascorbic acid, which dissolved in the network of thehydrated HA gel. Heating was not used to dissolve the AA in the gel.Propylene glycol, glycerol and ethyl alcohol were added and the mixturestirred at room temperature for approximately 1 hour. The Hylasomeincorporated with the above ingredients became clear in appearance. Tothis gel slurry was added Carbopol 940, and after uniform dispersement,the preservatives EDTA and phenonip were added, as well as natural lemonoil (water soluble) and TEA. The gel slurry remained transparent.Composition #2 was prepared following the same procedure, with theexception that mannitol was added to the gel slurry before Carbopol 940had been dispersed. Composition #2 was cloudy due to the presence ofmannitol, which did not fully dissolve in the gel slurry. Mannitol wasincluded in the composition in order to increase the stability of thegel slurry. The pH of both compositions was approximately 4.0.

Step #2: Stability Study of AA in the Gel Slurry (Compositions #1 and#2)

Concentration of AA (%) g/100 g Reduction of AA, % Composi- Composi-Composi- Composi- Time tion #1 tion #2 tion #1 tion #2 0 9.5 9.3 — — 1month 9.6 9.4 0 0 (no change) (no change) 12 months 8.6 8.5 10 9

The stability study was carried out at room temperature (20°-25° C.) andcold temperature (4°-7° C.). Samples were placed into 20 cc transparentglass containers with screw caps. The product occupied two-thirds of thevolume of the container. The rest (head-space) was air. Samples whichwere stored at room temperature were placed under artificial light, andthe stability was evaluated by measuring the concentration of AA using aspectrophotometric method, using a Perkin Elmer spectrometer, Model PELamda 2. Absorbance values were read at ?max243 (nm). All samples werediluted 10⁴ times using HCl to achieve a pH of approximately 2.0. Eachdata point in the table represents the average of a minimum of threevalues from samples of each composition. Absorbance reading weremeasured two times. Samples which were stored at cold temperaturesproduced absorbance values which were very close to those from thesamples stored at room temperature. Color change was also monitoredvisually, and were consistent with absorbance values. After 12 months ofstorage the color of the samples were yellow-amber, this color remainedstable for over 20 months.

Composition #3

Ingredients % (gm/100 gm) Ascorbic Acid 10 Glycerol 43 Mannitol 5Hylasome 22 Propylene Glycol 20

Reduction of AA concentration at different pH after 3 and 6 months ofstorage (including daylight). The pH was adjusted using TEA.

Reduction AA of AA Adjusted concentration, concentration, Original AAconc, % % % Composition AA Original After After After After Comp. #3conc, % pH 3 mo 6 mo 3 mo 6 mo 10.1 2.51 9.85 9.51 2.48 5.84 pH 3.0 9.679.48 9.25 1.94 4.34 pH 4.0 8.33 8.16 7.92 2.04 4.92 pH 5.0 6.43 6.3 6.182.02 3.89

The data presented in the preceding table shows that there is nocorrelation between AA stability and pH in compositions in which AA isstabilized by different polyhydroxy compounds.

Example 8

This example illustrates the preparation of a dermatological compositionusing Hylasome swollen with 50% trichloroacetic acid (TCA) as the activeingredient. To prepare 100 gm of a dermatological peeling composition,50 gm of TCA were added to 25 gm of water, 2.4 gm of glucose and 2.6 gmof glycerol and the mixture was stirred. To the mixture were added 20 gmof Hylasome (HA 1%) and it was stirred until fully swollen. The pH ofthe peeling composition was approximately 1.8-2.0.

Example 9

This example illustrates the preparation of the active ingredientHylasome swollen with a solution in water or saline of low molecularweight HA. To prepare 100 gm of Hylasome-HA gel, 99 gm of 0.1% HA(50,000 MW) were added to 1 gm of Hylasome (HA concentration 1%). Mixingwas continued until fully swollen. The total HA content was 0.11% andthe swelling ratio was 909. This composition can be used to control therelease of low molecular weight HA.

Example 10

This example illustrates the preparation of a composition with theactive ingredient Hylasome swollen with high molecular weight HA. Toprepare 100 gm of this composition 25 gm of water were added to 25 gm ofHylasome (HA 1%) and mixed until a uniform swollen gel was obtained witha HA concentration of 0.5%. To this gel 50 gm of 0.5% HA solution inwater (MW 500,000-6,000,000) were added and mixed until fully swollen.To 90 gm of equilibrated gel 10 gm of Centella Asiatica extract wereadded and stirred until uniform. This composition is used to treat theskin following other skin treatments such as skin peels anddermabrasion. The total HA concentration was 0.75%, the swelling ratiowas 133. This active ingredient may be used as described as a finalformulation, or as an ingredient in a formulation to control the releaseof low molecular weight actives into the skin.

Example 11

This example illustrates the preparation of a composition in whichHylasome (HA 1%) is incorporated with a hydrophobic material that is asunscreen (Parsol MCX).

Hylasome (HA 1%) was prepared according to Example 1. To Prepare 100 gmof the composition, 4.6 gm of lanoil AWS were added to 4.6 gm of VitaminE acetate (tocopheryl acetate) and 4.6 gm of Tween 80, and mixed untildissolved. Upon dissolution 41.4 gm of sunscreen Parsol MCX were addedand the mixture stirred until a uniform suspension formed. Then 18.4 gmof water were added and mixed, and 18.4 gm of Hyalsome (HA 1%) wereadded, and mixed until uniform. Then, 7.6 gm of noncrosslinkedhyaluronic acid (HA) were added and the mixture stirred. After mixing,0.4 g of phenonip preservative was added. The composition washomogenized according to the procedure described in Example 2. Theconcentration of sunscreen in this composition was 41.4%, theconcentration of Hylasome (HA 1%) was 18.4%, and the total HAconcentration was approximately 0.1% (1 mg/ml). To prepare 1 kg of thefinal formulation with sunblock properties, 900 g of the basicformulation which was prepared according to Example 2 were added to 100gm of Hylasome-sunscreen (Parsol MCX) suspension and thoroughly mixeduntil the consistency was that of a uniform cream. The final formulationhad an SPF of 19 (in vitro). The total concentration of sunscreen was5.04%, Hylasome (HA 1%) was 1.84% and the concentration of HA wasapproximately 0.12% (1.2 mg/ml). The Hyalsome-Parsol (MCX) compositionwas able to maintain the topical sunscreen for longer periods of time,and hence the potential to increase the duration of its effect (FIG. 1).There is also the potential to reduce the absorption of sunscreen intothe layers of the skin. Sunscreen which has been absorbed into the skinis no longer effective as a sunscreen, and also presents issues ofsafety due to systemic absorption. The resulting cream was stable andhad good hydrating properties, as well as an excellent feel on the skinand a pleasant smell due to the presence of natural oils and botanicalextracts.

Example 12

This example describes the preparation of a formulation with the activeingredient capsaicin and Hylasome.

Step 1: Preparation of a Complex of Capsaicin with Hylasome (HA 0.125%).

Hylasome with an HA concentration of 0.125% was prepared according toExample 2 with the exception that 1 part of Hylasome gel was added to 7parts of water, and phenonip was not added. The resultant gel was loadedwith capsaicin (Kalsec, 14% capsicum) at a ratio of gel:capsaicin of1.33 to 1. The gel was partially swollen with capsaicin.

Step 2: Preparation of the Basic Formulation with Cooling Effect.

This composition was prepared in separate stages as follows: Part A ofthe mixture was prepared by dispersing Carbopol 940 in water and thenstirring until uniform dispersion forms.

Formula % Part Ingredients by weight 1 A 1 Carbopol 940 0.48 2 water73.5 B 3 Cellosize polymer PCG-10 1% 7.68 4 Polyox 1% in water 0.34 5 HA1% in water 0.98 6 Ketrol 1% in water 4.8 7 Phosal 50 0.44 C 1 Robane0.14 2 Macadamia oil 0.48 3 Coconut oil 0.48 4 Olive oil 0.14 5 Cucumberextract 0.14 6 Chamomile extract 0.14 7 Calendula extract 0.48 8 AloeVera 0.14 9 Phenonip 0.75 10 Soy Protein 0.50 D 1 Menthol 5% in ethanol0.96 E 1 Triethanolamine 0.48 F 1 Lemon Natural Water Soluble 1.44 2Rose Natural water soluble 1.44 3 Dye - FD&C BLUE #1/ 0.07 FD&C YELLOW#6 9:1, 0.1% in water 2 1 Hylasome (HA 0.1%) 4.0

Parts B and C were prepared separately, and the three parts (A, B and C)were heated separately to 50-55° C. Then, Parts A, B and C were combinedand cooled to 30-35° C. Then Parts D, E, and F were added and mixedcarefully until a uniform gel-based cream formed. Part 2—Hylasome (HA0.1%) was then added.

Step 3: Preparation of the Final Formulation with Complex:Hylasome-Capsaicin.

To prepare 100 gm of the final formulation, 5 gm of TiO₂ were added to81.7 gm of the basic formulation (Step 2) and stirred well. Then, 13.3gm of the complex Hylasome (HA 0.125%)-capsaicin were added (Step 1).

Concentration of active ingredients: capsicum: 0.8%; HA: total HAconcentration was calculated based on dry polymer, 0.028%. When appliedto the skin a slight stinging sensation was observed; this reaction wastemporary and disappeared within minutes.

Example 13

This example illustrates the preparation of a formulation in whichHylasome (HA 1%) was incorporated with drugs such as an antibiotic(bacitracin), and a steroid (hydrocortisone or hydrocortisone acetate).

Step 1: Preparation of a Composition in which Hylasome (HA 1%) isIncorporated with Petrolatum.

To prepare 100 gm of this composition, first a mixture of 5 gm ofvitamin E acetate with 5 gm of Lanoil AWS and 5 gm of Tween 20 wasprepared. 45 gm of petrolatum were added to this composition and mixedcarefully. To this mixture were added 20 gm of water, mixed untiluniform suspension formed and then 20 gm of Hylasome (HA 1%) were added.

Step 2: Preparation of Hylasome-Drug Composition.

To prepare 25 gm of this composition, 0.45 gm of bacitracin and 1.25 gmof hydrocortisone (and/or hydrocortisone acetate) were added to 23.3 gmof Hylasome-petrolatum and carefully mixed.

Step 3: Preparation of the Final Formulation withHylasome-Petrolatum-Drugs.

To prepare 100 gm of the final formulation, a dermatological cream, 80gm of the formulation described in Example 2, Step 3, were added to 20gm of the composition described in Step 2 of this example. Theconcentration of the antibiotic in this cream was 0.36%, theconcentration of the cortisone was 1%. The concentration of thepetrolatum was 8.4%. The final dermatological composition possessedhighly hydrating, non-greasy properties due to the presence of Hylasome.This cream was effective in treating dry, cracked skin.

Example 14

This example illustrates the preparation of a composition in whichHylasome (HA 0.1%) which has been very highly homogenized to fineparticles into which a drug (povidone iodine) was incorporated. Tohylasome (HA 1%) prepared according to Example 2 (100 g) was added 1-12%by wt. of povidone iodine. This composition can be used as anantiseptic, antimicrobial, in the form of liquid drops for the treatmentof infections of the ear (otitis externa and otitis media), nose andthroat. The composition may also contain seabuckthorn oil (antiseptic)and rose oil for natural fragrance, and a pleasant smell to an otherwiseunpleasant smelling composition.

1. A topical composition comprising: a polysaccharide gel matrix; and asubstance, alone, or in combination with, an effective amount of atherapeutic agent deliverable to a skin surface, wherein the therapeuticagent is a salt, a solvate, a prodrug, or a derivative of thetherapeutic agent, and wherein the substance, alone or in combination,is entrapped within the matrix.
 2. A composition according to claim 1,wherein the polysaccharide is a cross-linked polymer of hyaluronic acid,a salt or a derivative of the polymer.
 3. A composition according toclaim 2, wherein the polymer is in an equilibrium form and is hylan B.4. A topical composition according to claim 2, wherein the polymer is ina non-equilibrium form and is hylan B.
 5. A composition according toclaim 1, wherein the substance is a hydrophilic organic ingredient.
 6. Acomposition according to claim 5, wherein the hydrophilic organicingredient is an α-, β-, γ- or δ-hydroxy acid, an α-amino acid, aβ-amino acid, a γ-amino acid, a δ-amino acid, a glycerin, a vitamin, avitamin derivative, a sugar, an essential oil, a water-soluble oil, afragrance, a natural polymer, a synthetic polymer, a cross-linkednatural polymer, a cross linked synthetic polymer, a peptide, a protein,a polysaccharide or a nucleic acid.
 7. A composition according to claim6 wherein the hydrophilic organic ingredient is an α-hydroxy acid,namely, lactic acid.
 8. A composition according to claim 1, wherein thesubstance is a hydrophobic organic ingredient.
 9. A compositionaccording to claim 8, wherein the hydrophobic organic ingredient is apetrolatum, a fatty acid, a fatty alcohol, an oil, a lipid or asilicone.
 10. A composition according to claim 1, wherein the substanceis an inorganic ingredient.
 11. A composition according to claim 10,wherein the inorganic ingredient is titanium dioxide, silica, calciumchloride, kaolin, talc or tantalum.
 12. A composition according to claim1, wherein the therapeutic agent is an anti-inflammatory agent, anantiseptic agent, an anti-infective agent, a mucous membrane agent, acleansing agent, a preservative, an astringent, an antihistamine, ananalgesic, an anesthetic, an acne medication, an antifungal agent, ananti-pruritic agent, a hormone, a growth factor, a moisturizing agent, asunscreen, a hyperpigmentation agent, an antioxidant or a nutritionalagent.
 13. A composition according to claim 12, wherein the nutritionalagent is a vitamin.
 14. A composition according to claim 12, wherein thesunscreen is Parsol MCX.
 15. A composition according to claim 12,wherein the therapeutic agent is an anti-inflammatory agent incombination with an anti-infective agent.
 16. A composition according toclaim 15, wherein the anti-inflammatory agent is a steroid agent.
 17. Acomposition according to claim 16, wherein the steroid agent ishydrocortisone, hydrocortisone acetate or prednisone.
 18. A compositionaccording to claim 15, wherein the anti-infective agent is anantibiotic.
 19. A composition according to claim 18, wherein theantibiotic is bacitracin.
 20. A composition according to claim 12,wherein the antiseptic agent is povidone iodine.